Specific Bacterial Proteins Research Articles

Myrtillin Prolongs Lifespan and Enhances Tolerance against Pathogenic Infections in Caenorhabditis elegans

BACKGROUND: Myrtillin is a pigment found abundantly in a variety of fruits and vegetables. As an anthocyanin compound, myrtillin exhibits antioxidant properties that can potentially counteract oxidative damage. OBJECTIVE: In this study, we focus on investigating the protective effects of myrtillin on Caenorhabditis elegans against pathogen-induced oxidative damage. Specifically, we explore the impact of myrtillin on the lifespan of C. elegans when exposed to different pathogens, namely Staphylococcus aureus, Enterococcus faecalis, Klebsiella pneumoniae, and Proteus mirabilis. MATERIALS AND METHODS: To conduct our experiments, C. elegans were administered varying concentrations of myrtillin and subsequently infected with each of the aforementioned pathogens. The nematodes’ lifespan was carefully monitored under these conditions. We also docked the three-dimensional structure of myrtillin to the crystal structure of different bacterial proteins. We visualized the crystal-docked structures with the lowest binding energy. RESULTS: Our findings revealed a significant increase in the lifespan of C. elegans when exposed to S. aureus, E. faecalis, K. pneumoniae, and P. mirabilis (P ≤ 0.05). It is worth noting that the effectiveness of myrtillin in extending lifespan varied among these pathogens. Notably, we observed that myrtillin exhibited low binding energies toward specific bacterial proteins, namely filamenting temperature-sensitive mutant Z of S. aureus and NAD+-dependent DNA ligase of E. faecalis. These interactions involved a combination of hydrogen bonds and hydrophobic interactions. In addition, myrtillin demonstrated the lowest binding energy with SHV-2 of K. pneumoniae and amidohydrolase of P. mirabilis, characterized by a higher number of hydrogen bonds. CONCLUSION: Based on our observations, we speculate that myrtillin possesses an additional protective mechanism against a range of pathogens, ultimately leading to an extended lifespan in C. elegans. This suggests the potential of myrtillin as a beneficial compound with diverse protective properties against various pathogens. Further research is required to elucidate the precise mechanisms underlying myrtillin’s effects and its potential application in promoting health and longevity.

Metabolic labeling probes for interrogation of the host-pathogen interaction.

Bacterial infections are still one of the leading causes of death worldwide; despite the near-ubiquitous availability of antibiotics. With antibiotic resistance on the rise, there is an urgent need for novel classes of antibiotic drugs. One particularly troublesome class of bacteria are those that have evolved highly efficacious mechanisms for surviving inside the host. These contribute to their virulence by immune evasion, and make them harder to treat with antibiotics due to their residence inside intracellular membrane-limited compartments. This has sparked the development of new chemical reporter molecules and bioorthogonal probes that can be metabolically incorporated into bacteria to provide insights into their activity status. In this review, we provide an overview of several classes of metabolic labeling probes capable of targeting either the peptidoglycan cell wall, the mycomembrane of mycobacteria and corynebacteria, or specific bacterial proteins. In addition, we highlight several important insights that have been made using these metabolic labeling probes.

Bacterial Protein Mimetic of Peptide Hormone as a New Class of Protein- based Drugs.

Specific peptide molecules classified as hormones, neuropeptides and cytokines are involved in intercellular signaling regulating various physiological processes in all organs and tissues. This justifies the peptidergic signaling as an attractive pharmacological target. Recently, a protein mimetic of a peptide hormone has been identified in Escherichia coli suggesting the potential use of specific bacterial proteins as a new type of peptide-like drugs. We review the scientific rational and technological approaches leading to the identification of the E. coli caseinolytic protease B (ClpB) homologue protein as a conformational mimetic of α-melanocyte-stimulating hormone (α-MSH), a melanocortin peptide critically involved in the regulation of energy homeostasis in humans and animals. Theoretical and experimental backgrounds for the validation of bacterial ClpB as a potential drug are discussed based on the known E. coli ClpB amino acid sequence homology with α-MSH. Using in silico analysis, we show that other protein sources containing similar to E. coli ClpB α-MSH-like epitopes with potential biological activity may exist in Enterobacteriaceae and in some Brassicaceae. Thus, the original approach leading to the identification of E. coli ClpB as an α-MSH mimetic protein can be applied for the identification of mimetic proteins of other peptide hormones and development of a new type of peptide-like protein-based drugs.

Dietary Nutrients, Proteomes, and Adhesion of Probiotic Lactobacilli to Mucin and Host Epithelial Cells

The key role of diet and environment in human health receives increasing attention. Thus functional foods, probiotics, prebiotics, and synbiotics with beneficial effects on health and ability to prevent diseases are in focus. The efficacy of probiotic bacteria has been connected with their adherence to the host epithelium and residence in the gut. Several in vitro techniques are available for analyzing bacterial interactions with mucin and intestinal cells, simulating adhesion to the host in vivo. Proteomics has monitored and identified proteins of probiotic bacteria showing differential abundance elicited in vitro by exposure to food components, including potential prebiotics (e.g., certain carbohydrates, and plant polyphenols). While adhesion of probiotic bacteria influenced by various environmental factors relevant to the gastrointestinal tract has been measured previously, this was rarely correlated with changes in the bacterial proteome induced by dietary nutrients. The present mini-review deals with effects of selected emerging prebiotics, food components and ingredients on the adhesion of probiotic lactobacilli to mucin and gut epithelial cells and concomitant abundancy changes of specific bacterial proteins. Applying this in vitro synbiotics-like approach enabled identification of moonlighting and other surface-located proteins of Lactobacillus acidophilus NCFM that are possibly associated with the adhesive mechanism.

Secretion Systems and Secreted Proteins in Gram-Negative Entomopathogenic Bacteria: Their Roles in Insect Virulence and Beyond.

Many Gram-negative bacteria have evolved insect pathogenic lifestyles. In all cases, the ability to cause disease in insects involves specific bacterial proteins exported either to the surface, the extracellular environment, or the cytoplasm of the host cell. They also have several distinct mechanisms for secreting such proteins. In this review, we summarize the major protein secretion systems and discuss examples of secreted proteins that contribute to the virulence of a variety of Gram-negative entomopathogenic bacteria, including Photorhabdus, Xenorhabdus, Serratia, Yersinia, and Pseudomonas species. We also briefly summarize two classes of exported protein complexes, the PVC-like elements, and the Tc toxin complexes that were first described in entomopathogenic bacteria.

Disparate subcellular location of putative sortase substrates in Clostridium difficile

Clostridium difficile is a gastrointestinal pathogen but how the bacterium colonises this niche is still little understood. Sortase enzymes covalently attach specific bacterial proteins to the peptidoglycan cell wall and are often involved in colonisation by pathogens. Here we show C. difficile proteins CD2537 and CD3392 are functional substrates of sortase SrtB. Through manipulation of the C-terminal regions of these proteins we show the SPKTG motif is essential for covalent attachment to the cell wall. Two additional putative substrates, CD0183 which contains an SPSTG motif, and CD2768 which contains an SPQTG motif, are not cleaved or anchored to the cell wall by sortase. Finally, using an in vivo asymmetric cleavage assay, we show that despite containing a conserved SPKTG motif, in the absence of SrtB these proteins are localised to disparate cellular compartments.

The Role of CagA in the Gastric Biology of Helicobacter pylori.

See related article by Blaser et al., [Cancer Res 1995;55:2111–5][1] . Visit the Cancer Research 75th Anniversary [timeline][2]. Helicobacter pylori (H. pylori) was discovered in 1983 by the Australian scientists Warren and Marshall as a gastric pathogen, causing peptic ulcer disease. In 2005,

NAIP proteins are required for cytosolic detection of specific bacterial ligands in vivo.

NLRs (nucleotide-binding domain [NBD] leucine-rich repeat [LRR]-containing proteins) exhibit diverse functions in innate and adaptive immunity. NAIPs (NLR family, apoptosis inhibitory proteins) are NLRs that appear to function as cytosolic immunoreceptors for specific bacterial proteins, including flagellin and the inner rod and needle proteins of bacterial type III secretion systems (T3SSs). Despite strong biochemical evidence implicating NAIPs in specific detection of bacterial ligands, genetic evidence has been lacking. Here we report the use of CRISPR/Cas9 to generate Naip1(-/-) and Naip2(-/-) mice, as well as Naip1-6(Δ/Δ) mice lacking all functional Naip genes. By challenging Naip1(-/-) or Naip2(-/-) mice with specific bacterial ligands in vivo, we demonstrate that Naip1 is uniquely required to detect T3SS needle protein and Naip2 is uniquely required to detect T3SS inner rod protein, but neither Naip1 nor Naip2 is required for detection of flagellin. Previously generated Naip5(-/-) mice retain some residual responsiveness to flagellin in vivo, whereas Naip1-6(Δ/Δ) mice fail to respond to cytosolic flagellin, consistent with previous biochemical data implicating NAIP6 in flagellin detection. Our results provide genetic evidence that specific NAIP proteins function to detect specific bacterial proteins in vivo.

A Molecular-Level Landscape of Diet-Gut Microbiome Interactions: Toward Dietary Interventions Targeting Bacterial Genes.

ABSTRACTAs diet is considered the major regulator of the gut ecosystem, the overall objective of this work was to demonstrate that a detailed knowledge of the phytochemical composition of food could add to our understanding of observed changes in functionality and activity of the gut microbiota. We used metatranscriptomic data from a human dietary intervention study to develop a network that consists of >400 compounds present in the administered plant-based diet linked to 609 microbial targets in the gut. Approximately 20% of the targeted bacterial proteins showed significant changes in their gene expression levels, while functional and topology analyses revealed that proteins in metabolic networks with high centrality are the most “vulnerable” targets. This global view and the mechanistic understanding of the associations between microbial gene expression and dietary molecules could be regarded as a promising methodological approach for targeting specific bacterial proteins that impact human health.

Cell-type specific control of immune responses mediated by inflammasomes in vivo (INM6P.333)

Abstract Upon pathogen recognition, certain cytosolic nucleotide-binding domain, leucine-rich repeat containing proteins (NLRs) form multi-protein complexes called inflammasomes, which activate CASPASE-1. CASPASE-1 initiates inflammation by processing IL-1beta and -18 into their active forms, and by inducing a rapid, lytic cell death called pyroptosis. The NLRC4 inflammasome responds to the cytosolic presence of specific bacterial proteins, such as flagellin, via NAIPs. A fundamental question is whether and how activation of innate immune responses triggers pathogen-specific adaptive immune responses. We seek to test the hypothesis that cell-type-specific activation of the inflammasome is sufficient to induce inflammation and adaptive immune responses in vivo. To test our hypothesis, we are using a novel ‘knock-in’ mouse strain. This mouse expresses a fusion protein, which selectively activates the NLRC4 inflammasome. Preliminary data indicate extensive NLRC4-dependent inflammatory disease in mice expressing the fusion protein in myeloid-derived cells.

Effectors, effectors et toujours des effectors

Effectors, effectors <i>et toujours des</i> effectors

An in vivo gene deletion system for determining temporal requirement of bacterial virulence factors

Analysis of phenotypes associated with specific mutants has been instrumental in determining the roles of a bacterial gene in a biological process. However, this technique does not allow one to address whether a specific gene or gene set is necessary to maintain such a process once it has been established. In the study of microbial pathogenesis, it is important but difficult to determine the temporal requirement of essential pathogenic determinants in the entire infection cycle. Here we report a Cre/loxP-based genetic system that allowed inducible deletion of specific bacterial genes after the pathogen had been phagocytosed by host cells. Using this system, we have examined the temporal requirement of the Dot/Icm type IV protein transporter of Legionella pneumophila during infection. We found that deletion of single essential dot/icm genes did not prevent the internalized bacteria from completing one cycle of intracellular replication. Further analyses indicate that the observed phenotypes were due to the high stability of the examined Dot/Icm protein. However, postinfection deletion within 8 h of the gene coding for the Dot/Icm substrate, SdhA, abolishes intracellular bacterial growth. This result indicates that the Dot/Icm transporter is important for intracellular bacterial growth after the initial biogenesis of the vacuole. Our study has provided a technical concept for analyzing the temporal requirement of specific bacterial proteins or protein complexes in infection or development.

Role of mineral specific bacterial proteins in selective flocculation and flotation

Extracellular bacterial protein (EBP) isolated from Paenibacillus polymyxa was fractionated through ammonium sulphate precipitation (ASP) method and characterized through SDS-PAGE. Role of such fractionated proteins in selective mineral separation was demonstrated through microbially-induced selective flotation and flocculation. Proper use of fractionated protein groups rendered pyrite and chalcopyrite hydrophilic while quartz, sphalerite and galena exhibited enhanced surface hydrophobicity after biotreatment. Sphalerite, galena and quartz could be selectively separated from pyrite–chalcopyrite through prior interaction with bacterial proteins.

Investigation of MALDI-TOF and FT-MS techniques for analysis of Escherichia coli whole cells.

Recently, it has been demonstrated that bacteria can be characterized using whole cells and matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). However, identification of specific bacterial proteins usually requires analysis of cellular fractions or purified extracts. Here, the first application of Fourier transform mass spectrometry (FTMS) to analysis of bacterial proteins directly from whole cells is reported. It is shown that accurate mass MALDI-FTMS can be used to characterize specific ribosomal proteins directly from Escherichia coli cells. High-accuracy mass measurements and high-resolution isotope profile data confirm posttranslational modifications proposed previously on the basis of low-resolution mass measurements. Seven ribosomal proteins from E. coli whole cells were observed with errors of less than 27 ppm. This was accomplished directly from whole cells without fractionation, concentration, or overt overexpression of characteristic cellular proteins. MALDI-FTMS also provided information regarding E. coli lipids in the low-mass region. Although ions with m/z values below 1000 have been observed by FTMS of whole cells, this represents the first report of detection of ions in the 5000 to 10,000 m/z range by MALDI-FTMS using whole cells.

IcsA heads to the pole

The localization of specific bacterial proteins to certain parts of the cell envelope can be visualized using fluorescence microscopy. These studies have shown that certain bacterial proteins have the capacity to be partitioned to specific areas of the cell where they perform their specific function. One such protein is the Shigella flexneri virulence factor IcsA, which is located toward one pole of the bacterium and requires this location to mediate intracellular motility. In a recent paper, Charles et al. 1 Charles M. et al. Polar targeting of Shigella virulence factor IcsA in Enterobacteriaceae and Vibrio. Proc. Natl. Acad. Sci. U. S. A. 2001; 98: 9871-9876 Crossref PubMed Scopus (87) Google Scholar used various IcsA–GFP fusion constructs to identify the IcsA polar-localization domain. Not only did the investigators find two distinct IcsA polar-targeting domains, but they also discovered evidence for a conserved polar-localization mechanism in species of Enterobacteriaceae and Vibrio.

A novel proline-rich protein, EspF, is secreted from enteropathogenic Escherichia coli via the type III export pathway

Enteropathogenic Escherichia coli (EPEC) cause a characteristic attaching and effacing (A/E) lesion in intestinal epithelial cells that is associated with the expression and export of specific bacterial proteins via a type III secretion pathway. These effector proteins and components of the type III export apparatus are encoded on a pathogenicity island known as the locus of enterocyte effacement (LEE). In this study, we describe a proline-rich protein, EspF, encoded by the LEE that is secreted by the EPEC type III secretion apparatus. Whereas an espF deletion mutant does not synthesize or secrete EspF, surprisingly it retains the ability to induce host signaling events, perform A/E activities, and invade host epithelial cells. Although these results do not indicate an obvious role for EspF in the formation of A/E lesions nor in the invasion of epithelial cells, they do not preclude a role played by EspF in other aspects of EPEC pathogenesis.

A novel proline-rich protein, EspF, is secreted from enteropathogenic Escherichia coli via the type III export pathway.

Enteropathogenic Escherichia coli (EPEC) cause a characteristic attaching and effacing (A/E) lesion in intestinal epithelial cells that is associated with the expression and export of specific bacterial proteins via a type III secretion pathway. These effector proteins and components of the type III export apparatus are encoded on a pathogenicity island known as the locus of enterocyte effacement (LEE). In this study, we describe a proline-rich protein, EspF, encoded by the LEE that is secreted by the EPEC type III secretion apparatus. Whereas an espF deletion mutant does not synthesize or secrete EspF, surprisingly it retains the ability to induce host signaling events, perform A/E activities, and invade host epithelial cells. Although these results do not indicate on obvious role for EspF in the formation of A/E lesions nor in the invasion of epithelial cells, they do not preclude a role played by EspF in other aspects of EPEC pathogenesis.

Nuclear import of Agrobacterium VirD2 and VirE2 proteins in maize and tobacco.

Previously, we have shown that Agrobacterium-plant cell transferred DNA (T-DNA) transport into the host cell nucleus is likely mediated by two specific bacterial proteins, VirD2 and VirE2. Here, we used these proteins to study molecular pathways of nuclear import. First, the role of VirE2 nuclear localization signals (NLSs) in the T-DNA transport pathway was examined by using tobacco plants transgenic for deletion mutants of VirE2. In these plants, the virulence of wild-type Agrobacterium was reduced possibly by competition for the cellular nuclear import machinery. Second, we analyzed the nuclear localization of VirE2 and VirD2 in the nonhost monocot maize. Part of the known recalcitrance of monocots to transformation by Agrobacterium could be due to a potential selectivity in nuclear import pathways in monocotyledonous and dicotyledonous plants. Nuclear transport of VirD2 and VirE2 in maize leaves and roots was compared to that in tobacco protoplasts and roots. Both proteins accumulated in maize leaf and tobacco protoplast nuclei as well as in nuclei of immature root cells. In contrast, VirD2 and VirE2 expressed in mature roots of maize and tobacco remained cytoplasmic. Point mutations of VirE2 nuclear localization signals, NSE 1 and NSE 2, also revealed that, in maize, the NSE 1 signal was mainly responsible for nuclear import; in contrast, both signals functioned independently in tobacco protoplasts.